Journal: bioRxiv

Article Title: Neuregulin-1 protects against respiratory viral induced mortality

doi: 10.1101/2023.05.10.540232

Figure Lengend Snippet: ( A ) CD11c + cells are increased in the lungs of atopic mice. Flow cytometry of lung cell suspension showing frequency (left) and cell number (right) at day 3 post inoculation (PI) high dose SeV ( B ) Adoptive transfer of CD11c + cells isolated from NA or atopic mice into naïve mice 24 h before inoculation with high dose SeV delays but does not prevent mortality; n=4 per group. ( C ) Transcriptomic (RNAseq) comparison between FACS isolated lung CD11c + cells from atopic and NA mice identifies several disparately expressed gene products, including Nrg1 (n=4 per group). Selected gene products shown. ( D ) NRG1 is markedly increased in atopic mouse lung (“tissue”), BAL, and supernatant from 1 x10 6 atopic lung CD11c + cells (“CD11c sup”) cultured for 24 h. ( E ) The scRNA (10x Platform) tSNE coordinates show CD14 + monocytes and dendritic cell (“DC”) sub-populations expressing NRG1 in peripheral blood cells of healthy human donors.*p<0.05, **p<0.01, ****p<0.0001

Article Snippet: Cell culture basal media was supplemented with recombinant human NRG1 alpha (cat#: NBP2-35093, Novus Bio) for hBEC or with mouse NRG1 for mTEC on day five and three before and at the time of infection with 4000 pfu of rgRSV or SeV-GFP.

Techniques: Flow Cytometry, Suspension, Adoptive Transfer Assay, Isolation, Comparison, Cell Culture, Expressing

Journal: bioRxiv

Article Title: Neuregulin-1 protects against respiratory viral induced mortality

doi: 10.1101/2023.05.10.540232

Figure Lengend Snippet: ( A ) NRG1(1ng to 1000ng) i.n. (in 30µL) given daily to naïve mice for 5d before inoculation with high dose SeV reduces viral mortality; n=4 per group (1ng, 10ng 1000ng & PBS), n=8 per group (100 ng and 500 ng). ( B ) Ratio of EBD in the BAL to that in the lung shows reduced EBD in NRG1 treated mice on day 8 PI SeV. n≥8 per group, median ± IQR shown, Mann-Whitney U test.

Article Snippet: Cell culture basal media was supplemented with recombinant human NRG1 alpha (cat#: NBP2-35093, Novus Bio) for hBEC or with mouse NRG1 for mTEC on day five and three before and at the time of infection with 4000 pfu of rgRSV or SeV-GFP.

Techniques: MANN-WHITNEY

Journal: bioRxiv

Article Title: Neuregulin-1 protects against respiratory viral induced mortality

doi: 10.1101/2023.05.10.540232

Figure Lengend Snippet: ( A ) Adding NRG1 to hBEC inoculated with rgRSV (left) and mTEC inoculated with GFP-SeV (right) reduces spread of infection. Representative images shown. ( B ) Quantification of (A) for rgRSV and hBEC and ( C ) for GFP-SeV and mTEC. GFP positive cells quantified by ImageJ. Representative images from ≥3 separate experiments. *p<0.05, **p<0.01. ( D ) Transcriptomic analysis of hBEC cultures treated with NRG1 (100 ng) on the basolateral side of the Transwell for 5 days and inoculated with RSV (4000 pfu). RNA was isolated 48 h PI RSV and qRT-PCR performed using a custom Prime PCR array plate: (i) Transcripts in which NRG1 treatment reduced gene expression from that seen in RSV infected cells. (ii) Transcripts with low level expression that show small but significant change in expression relative to naïve control with NRG1 alone or genes significantly increased with RSV but whose expression levels were not affected by NRG1. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001, n=3.

Article Snippet: Cell culture basal media was supplemented with recombinant human NRG1 alpha (cat#: NBP2-35093, Novus Bio) for hBEC or with mouse NRG1 for mTEC on day five and three before and at the time of infection with 4000 pfu of rgRSV or SeV-GFP.

Techniques: Infection, Isolation, Quantitative RT-PCR, Expressing, Control

Journal: bioRxiv

Article Title: Immune Checkpoint Molecules as Biomarkers of Staphylococcus aureus Bone Infection and Clinical Outcome

doi: 10.1101/2024.12.30.630837

Figure Lengend Snippet: (A) Immunofluorescent histochemistry analyses of tibia sections from uninfected and MRSA-infected humanized BLT mice 14 days post-op were performed with labeled antibodies against CD3, LAG-3, TIM-3, and PD-1 with DAPI counter stain, and representative images are shown at 4x. Note the increased numbers of T cells near the SAC (dashed yellow line) in the infected tibiae. (B) A multichromatic spectral flow cytometry analyses were performed on tibial bone marrow cells from uninfected and MRSA-infected BLT mice. Live human CD45+/CD3+/T cells and their subpopulations (CD4+, CD8+, Tregs) were analysed for immune checkpoint expression (LAG3, TIM-3, and PD-1) and proliferation (Ki67), and representative histograms are shown. Note the frequency of human CD3 + /CD4 + T cells expressing TIM-3, LAG3 & PD-1) in the cells from MRSA-infected bone marrow (n=4-8 mice, *p<0.05, t-test).

Article Snippet: Primary antibodies: The following antibodies were utilized for immunostaining: goat anti-CD3ε (clone M-20, sc-1127, RRID:AB_631128, Santa Cruz Biotechnology), mouse anti-PD-1 (10377-MM23, RRID:AB_2936309, Sino Biologicals), Rabbit anti-LAG3 (clone BLR027F, NBP2-76402, RRID:AB_3403543, Novus Biologicals), Mouse anti-TIM3/HAVCR2 (clone TIM3/4031, V8754-20UG, NSJ Bioreagents), Rabbit anti- S. aureus (PA1-7246, RRID:AB_561546, Thermo Fisher Scientific), and Mouse anti-CD66b (G10F5, NBP2-80664, RRID:AB_3096017, Novus Biologicals).

Techniques: Infection, Labeling, Staining, Flow Cytometry, Expressing

Journal: bioRxiv

Article Title: A hollow fiber membrane-based liver organoid-on-a-chip model for examining drug metabolism and transport

doi: 10.1101/2024.08.12.607504

Figure Lengend Snippet: (A) Representative images of intracellular fluorescence of calcein showing expanded monolayers after 7 days in EM. (B) Real-time qPCR analysis of the expression of ALB (albumin), CYP3A4 , CYP2C9 , CYP2D6 , CYP2B6 , UGT2A3 , SULT1A1 , ABCB1 (MDR1) and ABCC3 (MRP3) after 3 days in pre-DM and 7 days in DM. Data information: scale bars in A, 500 µm. Three independent replicates (donors; 2, 3, 5) with six technical replicates (monolayers) were examined. In B, the graphs show mean + SD (error bars). Three independent replicates (donors; 2, 3, 5; matched for all the conditions) in technical duplicates (wells) were examined.

Article Snippet: These included rabbit anti-ZO1 (1:50, Thermo Fisher Scientific, 40-2300), rabbit anti-OATP1B3 (1:50, Novus Biologicals, NBP1-80980), rabbit anti-OCT1 (1:50, Novus Biologicals, NBP1-59464), mouse anti-MDR1 (1:50, Santa Cruz Biotechnology, sc-55510), rabbit anti-MRP2 (1:50, Abcam, ab187644), rabbit anti-MRP3 (1:50, Novus Biologicals, NBP2-37923) and phalloidin (a toxin against F-actin) conjugated to Alexa Fluor 488 (1:400, Thermo Fisher Scientific, A12379).

Techniques: Fluorescence, Expressing

Journal: bioRxiv

Article Title: A hollow fiber membrane-based liver organoid-on-a-chip model for examining drug metabolism and transport

doi: 10.1101/2024.08.12.607504

Figure Lengend Snippet: (A) A representative whole-mount confocal immunofluorescence microscopy image of OATP1B3, OCT1, MDR1, MRP2 and MRP3 in a dorsal section of a HFM. (B) A representative confocal immunofluorescence microscopy image for the basolateral localization of NTCP in cell membranes of a monolayer in an anterior section of a HFM. (C) Representative brightfield images and images of intracellular fluorescence of calcein of monolayers treated with efflux inhibitors versus DMSO (control). (D) The quantification of calcein fluorescence. Data information: scale bars in A-C, 100 µm. In A and B, nuclei were counterstained with DAPI. In D, the graph shows mean ± SD (error bars). In A and B, three independent replicates (donors; 2, 3, 5) with three technical replicates (monolayers) were examined. In C and D, one independent replicate (donor, 5) with 40 technical replicates (fields) from two monolayers (20 replicates each) was examined.

Article Snippet: These included rabbit anti-ZO1 (1:50, Thermo Fisher Scientific, 40-2300), rabbit anti-OATP1B3 (1:50, Novus Biologicals, NBP1-80980), rabbit anti-OCT1 (1:50, Novus Biologicals, NBP1-59464), mouse anti-MDR1 (1:50, Santa Cruz Biotechnology, sc-55510), rabbit anti-MRP2 (1:50, Abcam, ab187644), rabbit anti-MRP3 (1:50, Novus Biologicals, NBP2-37923) and phalloidin (a toxin against F-actin) conjugated to Alexa Fluor 488 (1:400, Thermo Fisher Scientific, A12379).

Techniques: Immunofluorescence, Microscopy, Fluorescence, Control

Journal: iScience

Article Title: Spatiotemporally organized immunomodulatory response to SARS-CoV-2 virus in primary human broncho-alveolar epithelia

doi: 10.1016/j.isci.2023.107374

Figure Lengend Snippet:

Article Snippet: Anti-cytokeratin 5 AF594 (Polyclonal) , Novus , CatNBP2-61931AF594 RRID: AB_2933967.

Techniques: Bacteria, Virus, Variant Assay, Recombinant, Red Blood Cell Lysis, Software, Membrane, Cell Culture

Journal: Biochimica et biophysica acta. Molecular basis of disease

Article Title: Sesn3 deficiency promotes carcinogen-induced hepatocellular carcinoma via regulation of the hedgehog pathway

doi: 10.1016/j.bbadis.2019.07.011

Figure Lengend Snippet: Representative immunofluorescence analysis of Acta2 (α-SMA), Col1, and Col3 (A). Analysis of fluorescence intensity for Acta2, Col1, and Col3 (B). Hepatic mRNA levels of Pdgfrb, Col1a1, and Acta2 (C). Immunoblot analysis of hepatic Acta2 and Pdgfrb (D). Representative immunofluorescence analysis of hepatic AFP, CK-19, Arg-1, Cd133, and Cd44 (E). Analysis of fluorescence intensity of hepatic AFP, CK-19, Argl, Cd133, and Cd44 (F). Data are expressed as mean ± SEM (n = 4 to 6 each group). #p < 0.05 and ##p < 0.01 for KO vs. WT. Immunofluorescence image analyses were performed using a fluorescence microscope (ZEISS, x200 magnification).

Article Snippet: Immunofluorescence (IF) analysis was performed for detecting general HCC markers including Arginase-1 (Arg1, rabbit mAb, Cat. No. 93668S, Cell Signaling Technology), alpha-fetoprotein (AFP, mouse mAb, sc-8399, Santa Cruz Biotechnology), and cytokeratin 19 (CK-19, rat mAb, TROMA-III, DSHB), ECM molecules such as Collagen 1 and 3 (Col1, rabbit anti-Collagen I antibody, #ab21286; Col3, rabbit anti-Collagen III antibody, #ab7778, Abcam), and alpha-smooth muscle actin (α-SMA, rabbit anti-αSMA antibody, #ab5694, Abcam), and cancer stem cell markers like CD44 (mouse anti-CD44 antibody, #sc-7297, Santa Cruz Biotechnology) and CD 133 (rabbit anti-CD133 antibody, #NBP244250, Novus Biologicals).

Techniques: Immunofluorescence, Fluorescence, Western Blot, Microscopy

Journal: Biochimica et biophysica acta. Molecular basis of disease

Article Title: Sesn3 deficiency promotes carcinogen-induced hepatocellular carcinoma via regulation of the hedgehog pathway

doi: 10.1016/j.bbadis.2019.07.011

Figure Lengend Snippet: Differentially expressed genes (DEGs) from the RNA-seq analysis of WT and Sens3-KO livers were presented by the volcano plot (A). A selected list of DEGs were presented in the table (B). Top 10 biological processes were over-represented in the significantly upregulated (C) and downregulated (D) genes. Hepatic Cd44 and Cd133 mRNA analysis by real-time qPCR (E). Immunoblot analysis of cancer signaling-related proteins in the liver of WT and Sesn3 KO mice (F). Co-IP analysis of Sesn3 and Gli2 interaction in HEK 293T cells (G). Data are expressed as mean ± SEM (n=4 for the immunoblot analysis, n=10 for ELISA, n=6 for qPCR analysis, respectively). #p < 0.05 and ##p < 0.01 for KO vs. WT.

Article Snippet: Immunofluorescence (IF) analysis was performed for detecting general HCC markers including Arginase-1 (Arg1, rabbit mAb, Cat. No. 93668S, Cell Signaling Technology), alpha-fetoprotein (AFP, mouse mAb, sc-8399, Santa Cruz Biotechnology), and cytokeratin 19 (CK-19, rat mAb, TROMA-III, DSHB), ECM molecules such as Collagen 1 and 3 (Col1, rabbit anti-Collagen I antibody, #ab21286; Col3, rabbit anti-Collagen III antibody, #ab7778, Abcam), and alpha-smooth muscle actin (α-SMA, rabbit anti-αSMA antibody, #ab5694, Abcam), and cancer stem cell markers like CD44 (mouse anti-CD44 antibody, #sc-7297, Santa Cruz Biotechnology) and CD 133 (rabbit anti-CD133 antibody, #NBP244250, Novus Biologicals).

Techniques: RNA Sequencing Assay, Western Blot, Co-Immunoprecipitation Assay, Enzyme-linked Immunosorbent Assay